title: MicroRNA-561 promotes acetaminophen-induced hepatotoxicity in HepG2 cells and primary human hepatocytes through downregulation of the nuclear receptor corepressor dosage-sensitive sex-reversal adrenal hypoplasia congenital critical region on the X chromosome, gene 1 (DAX-1).
abstract: One of the major mechanisms involved in acetaminophen (APAP)-induced hepatotoxicity is hepatocyte nuclear factor 4α (HNF4α)-mediated activation of pregnane X receptor (PXR) and constitutive androstane receptor (CAR). In the present study, we investigated the role of miR-561 and its target gene DAX-1 encoding a corepressor of HNF4α in the process of APAP-induced hepatotoxicity. We used both human hepatocellular liver carcinoma cell line (HepG2) cells and primary human hepatocytes in this study and monitored the levels of reactive oxygen species, lactate dehydrogenase, and glutathione. Our bioinformatics study suggests an association between miR-561 and DAX-1, but not HNF4α. Treatment of HepG2 cells with APAP significantly reduced the expression of DAX-1 in a concentration-dependent manner. miR-561 was induced by APAP treatment in HepG2 cells. Transfection of HepG2 cells with an miR-561 mimic exacerbated APAP-induced hepatotoxicity. HNF4α is physically associated with DAX-1 in HepG2 cells. A decreased protein level of DAX-1 by APAP treatment was also enhanced by miR-561 mimic transfection in HepG2 cells and primary human hepatocytes. The basal and APAP-induced expression of PXR and CAR was enhanced by miR-561 mimic transfection; however, transfection of HepG2 cells or primary human hepatocytes with a miR-561 inhibitor or DAX-1 small interfering RNA reversed these effects. Additionally, the chromatin immunoprecipitation assay revealed that recruitment of DAX-1 onto the PXR promoter was inversely correlated with the recruitment of peroxisome proliferator-activated receptor-α coactivator-1α and HNF4α on APAP treatment. These results indicate that miR-561 worsens APAP-induced hepatotoxicity via inhibition of DAX-1 and consequent transactivation of nuclear receptors.
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, Florida (M.L., Z.W.Z., S.F.Z.); Department of Hepatobiliary Surgery, General Hospital of Ningxia Medical University, Yinchuan City, Ningxia Hui Autonomous Region, China (Y.Y.); Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center, Guiyang Medical University, Guiyang, Guizhou, China (Z.X.H.); Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, Utah (T.Y.); Nanobiotechnology Center and Markey Cancer Center, College of Pharmacy, University of Kentucky, Lexington, Kentucky (P.G.); Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, China (X.Z.).
authors.forename: Minghua
authors.initials: M
authors.lastname: Li
authors.forename: Yinxue
authors.initials: Y
authors.lastname: Yang
authors.forename: Zhi-Xu
authors.initials: ZX
authors.lastname: He
authors.forename: Zhi-Wei
authors.initials: ZW
authors.lastname: Zhou
authors.forename: Tianxin
authors.initials: T
authors.lastname: Yang
authors.forename: Peixuan
authors.initials: P
authors.lastname: Guo
authors.forename: Xueji
authors.initials: X
authors.lastname: Zhang
authors.forename: Shu-Feng
authors.initials: SF
authors.lastname: Zhou
mesh_terms: D000082:Acetaminophen / Q000009:adverse effects*; D000312:Adrenal Hyperplasia, Congenital / Q000235:genetics / Q000378:metabolism; D000309:Adrenal Insufficiency; D000975:Antioxidants / Q000378:metabolism; D045744:Cell Line, Tumor; D056486:Chemical and Drug Induced Liver Injury / Q000235:genetics* / Q000378:metabolism; D056970:Co-Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D000090702:Constitutive Androstane Receptor; D057137:DAX-1 Orphan Nuclear Receptor / Q000235:genetics* / Q000378:metabolism; D015536:Down-Regulation / Q000235:genetics*; D040181:Genetic Diseases, X-Linked / Q000235:genetics / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D006059:Gonadal Dysgenesis / Q000235:genetics / Q000378:metabolism; D056945:Hep G2 Cells; D051557:Hepatocyte Nuclear Factor 4 / Q000235:genetics / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D000075262:Hypoadrenocorticism, Familial; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics*; D000071248:Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; D000077297:Pregnane X Receptor; D011401:Promoter Regions, Genetic / Q000235:genetics; D017382:Reactive Oxygen Species / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism; D011987:Receptors, Steroid / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000235:genetics
chemical_list: D000975:Antioxidants; D056970:Co-Repressor Proteins; D000090702:Constitutive Androstane Receptor; D057137:DAX-1 Orphan Nuclear Receptor; C496592:HNF4A protein, human; D051557:Hepatocyte Nuclear Factor 4; D035683:MicroRNAs; C539965:NR0B1 protein, human; C486497:PPARGC1A protein, human; D000071248:Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; D000077297:Pregnane X Receptor; D017382:Reactive Oxygen Species; D018160:Receptors, Cytoplasmic and Nuclear; D011987:Receptors, Steroid; D014157:Transcription Factors; D000082:Acetaminophen; D007770:L-Lactate Dehydrogenase; D005978:Glutathione